Polymer field-effect transistors (PFETs) are among the most fascinating electronic devices because of their attractive properties, such as durability, environmental friendliness, and solution processability. To date, PFETs have been employed for the development of flexible displays, radio-frequency identification tags, flexible non-volatile memories, among others. Moreover, due to the above advantages, PFETs can be applied to disposable on-site analytical devices. In that regard, we have developed extended-gate type PFETs with molecular recognition biomaterials for protein sensing in aqueous media. The fabricated PFETs were used to successfully detect glycoproteins (immunoglobulin A, immunoglobulin G, and chromogranin A) without any complicated labeling processes. Since our proposed immunoassay, which is performed on the basis of extended-gate type PFETs, is rapid and easy-to-use, PFET will be an attractive platform for on-site monitoring devices in healthcare applications in the near future.